1
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:
:


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António G. Brito, João Peixoto, José M. Oliveira, José A. Oliveira,
Cristina Costa, Regina Nogueira, and Ana Rodrigues
*

1. INTRODUCTION
Environmental issues are a critical factor for the today industry competitiveness.
Indeed, the society and the individual consumers could set a common framework for
companies’ commitment and engagement regarding environment protection. Redesign
the process, recover by-products or reuse effluents are some of the possible actions
towards an eco-efficient strategy. Nevertheless, a point remains crucial in such mission:
the ability to defend natural ecosystems from polluted wastewaters. For such purpose, a
wastewater treatment plant that maximizes removal efficiency and minimizes investment

and operation costs is a key factor.
Brewery and winery are traditional industries with an important economic value in
the agro-food sector. In 2003, the total beer production in the European Union (18
countries) was 344
x 10
5
m
3
, being recorded around 1800 breweries with 110 thousand
employees. If Norway, Switzerland and Turkey are also included, those numbers rise up
to 358 x 10
5
m
3
, 1839 units and 117 thousand, respectively. The excise revenue from beer
industry in all these countries reaches over 8800 x 10
6
€ (The Brewers of Europe, 2004).
The worldwide wine production is 261 x 10
5
m
3
(data from 2002), of which 69 %
from Europe, 18 % from America, 5 % from Asia, 4 % from Africa and 4 % from
Oceania. The worldwide wine consumption (2002) is 228 x 10
5
m
3
, distributed by Europe
(68 %), America (20 %), Asia (7 %), Africa (3 %) and Oceania (2 %) (OIV, 2002).

This chapter intends to present some key points on design and operation in
wastewater treatment of brewery and winery industries. Therefore, an introduction of the
industrial processes is first presented and then wastewater characteristics and treatment
processes are discussed. Finally, the experience of a collaborative effort between


*
António G. Brito, João Peixoto, José M. Oliveira, Regina Nogueira, and Ana Rodrigues, University of Minho,
School of Engineering – Center of Biological Engineering, Campus de Gualtar, 4710-057 Braga, Portugal.
José A. Oliveira, Adega Cooperativa de Ponte da Barca, Lugar de Agrelos, 4980-601 Ponte da Barca,
Portugal. Cristina Costa, Unicer SA, Leça do Balio, Matosinhos, 4466-955 S. Mamede de Infesta, Portugal.
2 A. G. BRITO ET AL
University of Minho and two industrial companies, Unicer SA and ACPB (Adega
Cooperativa de Ponte da Barca) is presented in order to address some practical problems
of wastewater systems design and operation. Unicer SA and ACPB are very important
players in their field of activity: Unicer has the major share of the beer market in Portugal
and ACPB is a very well known producer of wine with appellation of origin Vinho Verde.
2. BREWERY AND WINERY INDUSTRIES: AN OVERVIEW
2.1. Brewing Processes
Beer is a soft drink obtained through alcoholic fermentation, using selected yeasts of
the genera Saccharomyces, of wort prepared from malt cereals, mainly barley, and other
amylaceous or sugar-based raw materials, to which were added hop flowers, or their
derivatives, and adequate water. Figure 1 shows a typical technological process.
MALTING
MASHING
WORT BOILING
HOPS
YEAST
MILLING
(CORN GRITZ, BARLEY,

RICE, WHEAT; ENZYMES;
SUGAR, SUGAR SYRUPS)
WORT FILTRATION
FERMENTATION
BY-PRODUCTS
(SPENT GRAINS)
MATURATION
STABILIZATION
CLARIFICATION
PACKAGING
SEDIMENT REMOVAL
(TRUB)
WASTEWATER
SOLIDS
WASTEWATER
SOLIDS
O
2
WATER
FINING AGENTS
ANTI-OXIDISING AGENTS
KIESELGUHR
BY-PRODUCTS
(SURPLUS YEAST)
BARLEY
WATER
BREWHOUSE OPERATIONS

Figure 1. Technological process in breweries (adapted from Unicer SA and Varnam and Sutherland, 1994).


BREWERY AND WINERY 3

A mass balance is depicted in Figure 2, which represents water and energy inputs,
and also the outputs respecting residues and sub-products, liquid effluents and air
emissions. Residues similar to urban residues, simple industrial residues, glass, paper,
cardboard, plastic, oils, wood, biological sludge, green residues, etc. are classified as
solid wastes; surplus yeast and spent grains are considered sub-products. Brewer’s spent
grains are generally used for the production of low value composts, livestock feed or
disposed of in landfill as waste (Jay et al., 2004). Alternatively, the spent grains can be
hydrolyzed for the production of xylo-oligosaccharides (probiotic effect), xylitol
(sweetener), or pentose-rich culture media (Carvalheiro et al., 2004 and 2005; Duarte et
al, 2004).
2.2. Winemaking Processes
Wine is the product obtained from the total or partial alcoholic fermentation of fresh
grapes, whether or not crushed, or of grape must. Producing wine requires the implemen-
tation of a biotechnological sequence involving several unit operations. Although some
few products are added to the must and/or wine, several residues are rejected, either as
liquid or solid wastes. White wine is normally produced by the fermentation of a clarified
must, which is obtained after grape stem removal, pressing of the resulted grape berries
and subsequent clarification. The production of red wines is usually conducted in non-
clarified musts, prepared after grape stem removal and crushing of grape clusters. Musts
can also be fermented in the presence of grape stems. After fermentation, wines must be
clarified and stabilized, chemically and microbiologically, before bottling. Figure 3
shows a schematic process, applied at ACPB, to produce wines (Vinho Verde). These
wines follow the ordinary winemaking process, but ageing is avoided, in order to
preserve the original freshness and fruity characteristics.

Water
4.87 m
3

/m
3
Beer
Production
Gas emissions
“greenhouse effect”

130.5 kg/m
3


SOLIDS
Electric energy
126.9 kWh/m
3

Thermal energy
1.13 GJ/m
3


Fossil Fuels
41.7 kg/m
3

Acidifying
emissions
1.1 kg/m
3


Wastewaters
3.3 m
3
/m
3
COD = 13.2 kg/m
3

Solid Wastes:

51.2 kg/m
3

Valorization index = 93 %
Sub-products:

143.6 kg/m
3

Valorization index = 100 %

Figure 2. Mass balance applied to Unicer SA breweries representing specific values, i. e., values per cubic
meter of produced beer (Unicer SA, 2005).
4 A. G. BRITO ET AL
GRAPE RECEPTION
(CLARIFICATION)
SO
2
YEAST
DESTEMMING + CRUSHING

FERMENTATION
TRANSFERS
LEES
WASTEWATER
CONSERVATION
FINING
FILTRATION
BOTTLING
TARTRATES
RESIDUES
SEDIMENTS
WASTEWATER
GRAPE STEMS
WASTEWATER
LEES + SEEDS
WASTEWATER
TARTRATES
WASTEWATER
SO
2
COLD STABILIZATION
SO
2
SO
2
POTASSIUM BICARBONATE
FINING AGENTS
KIESELGUHR
POTASSIUM BITARTRATE
GUM ARABIC

CO
2
WASTEWATER
(PRESSING)
SKINS + SEEDS
WASTEWATER

Figure 3. Technological process adopted at ACPB wine-cellar.
Wineries, distilleries and other grape processing industries annually generate large
volumes of wastewater. This mainly originates from various washing operations during
the crushing and pressing of grapes, as well as rinsing of fermentation tanks, barrels and
other equipment or surfaces (Petruccioli et al., 2000). Over the year, volumes and
pollution loads greatly vary in relation to the working period (vintage, racking, bottling)
and to the winemaking technologies used, e. g., in the production of red, white and
special wines (Rochard, 1995; Anon, 1996).
A mass balance of wine production is depicted in Figure 4, which represents water
and energy inputs, and also the outputs respecting residues and sub-products, as well as
liquid effluents. Simple municipal and some industrial residues (glass, paper, cardboard,
plastic, wood and filtration earths) but also yeasts, grape stems, pomace and lees should
be recycled and valorized whenever possible.
BREWERY AND WINERY 5



Figure 4. Mass balance applied to ACPB winery representing specific values, i. e., values per cubic meter of
produced wine (2004). Losses of water by evaporation were neglected.
Yeasts cannot be used in animal dietary because they have high contents of
polyphenols and may contain some residues coming from treatments; they can only be
composted with pomace. However, pomace, seeds, lees, effluents resulting from tartar
removal and wine rests can be valorized to produce compounds with adding value like

alimentary colorant E163, alimentary oil, tartaric acid, 1,3-propanediol and dihydroxy-
acetone (Bourzeix et al., 1998). On the other hand, the grape stems can be composted, the
final compost being used as organic soil amendment and the grape pomace can be sold to
distilleries.
3. WASTEWATER TREATMENT
3.1. Brewery Industry
3.1.1. Wastewater Characterization
The composition of brewing effluents can fluctuate significantly as it depends on
various processes that take place within the brewery, but the amount of wastewater
produced depends on the water consumption during the process. In general, water
consumption per volume of produced beer attain 4.7 m
3
/m
3
(Carlsberg, 2005) but it
should be pointed that the wastewater to beer ratio is often 1.2 m
3
/m
3
to 2 m
3
/ m
3
less
because part of the water is disposed off with by-products and lost by evaporation
(Drissen and Vereijken, 2003).
Organic components in brewery effluent are generally easily biodegradable and
mainly consist of sugars, soluble starch, ethanol, volatile fatty acids, etc., leading to a
Water
9.25 m

3
/m
3
Wine
Production
Electric energy
159.6 kWh/m
3

SOLIDS
Wastewaters
9.25 m
3
/m
3

Solid wastes: 27.4 kg/m
3

Valorization index = 43 %
Sub-products: 406.3 kg/m
3

Valorization index = 100
6 A. G. BRITO ET AL
BOD/COD
a
ratio of 0.6 to 0.7. The effluent solids consist of spent grains, kieselguhr,
waste yeast and “hot” trub. The pH levels are determined by the amount and the type of
chemicals used at the CIP (clean in place) units (e.g. caustic soda, phosphoric acid, nitric

acid). Nitrogen
b
and phosphorous levels are mainly depending on the handling of raw
material and the amount of spent yeast present in the effluent. High phosphorous levels
can also result from the chemicals used in the CIP unit. Table 1 summarizes some of the
most important environmental parameters.
Table 1. Characteristics of some industrial brewery effluents including Unicer’s
Parameter / benchmark
Brewery effluent composition
per unit
Unicer
Typical
a
Opaque beer
b

COD (mg/L) 800 – 3 500 2 000 – 6 000 8 240 – 20 000
BOD (mg/L) 520 – 2 300 1 200 – 3 600
TSS
c
(mg/L) 200 – 1 000 2 901 – 3 000
TS
c
(mg/L) 5 100 – 8 750
T
o
C 30 – 35 18 – 20 25 – 35
pH 6.5 – 7.9 4.5 – 12 3.3 – 6.3
Nitrogen (mg/L) 12 – 31 25 – 80 0.0196 – 0.0336
Phosphorous (mg/L) 9 – 15 10 – 50 16 – 24

(Water/Beer) (m
3
/m
3
) 4.87
(Liquid effluent/Beer) (m
3
/m
3
) 3.3 2 – 8
(COD/Beer) (kg/m
3
) 13.2 5 – 30
(BOD/Beer) (kg/m
3
) 2 – 20
(TSS/Beer) (kg/m
3
) 1 – 5
a
Driessen and Vereijken (2003).
b
Parawira et al., (2005)
c
TS, TSS – Total solids, total suspended solids.
3.1.2. Treatment Processes
Different environmental and socio-economics criteria can be considered when
deciding on a wastewater treatment plant for a brewery industry. The aim is to select a
process that is flexible enough to cope with large fluctuations in organic load and
characteristics of such wastewaters, while keeping capital and operating costs as low as

possible. Because organic matter concentration in brewery effluent is significant, a high
input of energy for aeration is required. Another factor is the amount of waste sludge
generated from aerobic metabolism, which also needs to be handled and disposed of.
Both increase the cost of operation of the treatment system. Therefore, anaerobic
processes are preferred for the purpose of brewery wastewaters pre-treatment because
energy is saved and sludge disposal costs are minimized. When discharging into surface


a
BOD – Biochemical oxygen demand – and COD – Chemical oxygen demand – (mass of O
2
per volume).
b
N – Nitrogen mass concentration (mass of N per volume). NO
3
–
-N, NO
2
–
-N, NH
4
+
-N – Nitrate, nitrite, and
ammonia mass concentration as mass of N per volume.
BREWERY AND WINERY 7

water bodies, anaerobic pre-treatment combined with subsequent aerobic post-treatment
for organic
or nutrient removal is considered to be the best solution (Rodrigues et al.,
2001; Nogueira et al., 2002).

Several types of anaerobic reactors can be applied to brewery wastewater treatment.
However, the Upflow Anaerobic Sludge Blanket (UASB) reactor clearly accounts for the
most usual full-scale systems (Batston et al., 2004; Parawira et al., 2005). The upflow
mode of operation induces the development of a characteristic biological self-aggregation
process without addition of support material. The resulting biofilm structure is usually
denominated “granules” and is the main factor for their high biomass concentration and
biological activity (Brito et al., 1997a). The Expanded Granular Sludge Bed (EGSB)
reactor is a tower reactor using granular anaerobic sludge, identical to UASB reactors,
built with tank heights of 12 m to 16 m. The Internal Circulation (IC) reactor also uses
granular anaerobic sludge and is built with higher tank heights (up to 24 m). Whereas the
EGSB and UASB reactors separate the biomass, biogas and wastewater in a 1-step three-
phase-separator located in top of the reactor, the IC reactor is a 2-staged UASB reactor
design. The lower UASB receives extra mixing by an internal circulation, driven by its
own gas production. While the first separator removes most of the biogas, turbulence is
significantly reduced, allowing the second separator effectively separating the anaerobic
sludge from the wastewater. The loading rate of the IC reactor, as COD, is typically twice
as high as the UASB reactor (15 kg m
–3
d
–1
to 30 kg m
–3
d
–1
). Another positive factor
resulting from the applied high hydraulic upflow velocities is the selective washout of
brewery solids, like kieselguhr, trub and yeast.
In order to meet stringent requirement of surface water quality, an aerobic polishing
step is necessary after the anaerobic pre-treatment. Sequencing batch reactors (SBR) are
well suited for such purpose (Brito et al., 1997b; Rodrigues et al., 2004). The SBR is a

periodic process that performs multiple biological reactions in non steady-state
conditions. Biomass retention throughout the introduction of a decanting step and the
ease of automation are additional advantages for using SBR technology (Rodrigues et al.,
1998). Nevertheless, some other interesting experiences regarding aerobic processes can
be named. Selected examples are jet loop reactors (Bloor et al., 1995), fluidised bed
bioreactor (Ochieng et al., 2002) and membrane bioreactors (Cornelissen et al., 2002). It
should be noted that membrane bioreactors deserve a special attention within the brewing
industry. Their market share can increase in the next few years, including in the anaerobic
concept (Ince et al., 2000).
3.2. Winery Industry
3.2.1. Wastewater Characterization
Winemaking is seasonal with high activity in autumn (at north hemisphere), which
corresponds to vintages and fermentations, a notoriously less important activity in spring
on the occasion of transfers (racking period) and filtrations, and a weak activity during
winter and summer. Winery effluents contain four types of principal pollutants:
• Sub-product residues – stems, seeds, skins, lees, sludge, tartar, etc.;
• Loss of brut products – musts and wines occurred by accidental losses and
during washings;
• Products used to wine treatments – fining agents, filtration earths, etc.;
8 A. G. BRITO ET AL
• Cleaning and disinfection products, used to wash materials and soils.
Musts and wines constituents are present in wastewaters, in variable proportions:
sugars, ethanol, esters, glycerol, organic acids (e.g., citric, tartaric, malic, lactic, acetic),
phenolic compounds (coloring matter and tannins) and a numerous population of bacteria
and yeasts. They are easily biodegradable elements, except for polyphenols (60 mg/L to
225 mg/L) which make this biodegradation more difficult and requiring an adapted flora.
Effluents have a pronounced demand in nitrogen and phosphorous, with a BOD
5
/N/P
relation often near 100/1/0.3 (Torrijos and Moletta, 1998). Additionally, effluents have a

daily great variability, in both quantity and quality, making evaluation of daily pollution
complex. Generally, the production of 1 m
3
of wine generates a pollution load equivalent
to 100 persons. The pH is usually acidic but, punctually, it may display basic values, on
the occasion of the cleaning operations (with alkaline products and organochlorides) and
on the occasion of chemical detartaration.
Rejected volumes per volume of produced wine vary from one wine cellar to
another, with extreme values comprised between 0.1 m
3
/m
3
and 2.4 m
3
/m
3
. For the ratio
of water consumption to produced wine, 1.0 m
3
/m
3
is the rule of thumb, while Pévost and
Gouzenes (2003) refer to values between 0.3 m
3
/m
3
and 2.5 m
3
/m
3

. Table 2 shows some
examples of winery effluents main characteristics. Washing operations carried out during
different winemaking steps, which are at the origin of the rejection of fully charged
wastewaters, can be distributed as follow:
– During vintage preparation – washing and disinfection of materials;
– During grape reception – washing of reception materials (hoppers, destemmers,
crushers, presses, dejuicers, conveyors and transport pumps); cleaning the floors, with or
without addition of cleaning products;
– During vinifications – rinsing of fermentation and clarification vats; cleaning the
floors, with or without addition of cleaning products;
– During transfers – rinsing vats after transfers; cleaning the floors, with or without
addition of cleaning products;
– During filtrations – rinsing kieselguhr and earth filters.
Table 2. Examples of effluent composition (mean or range values) of four different
wineries, including that of ACPB

Wine cellar
a


ACPB A
b
B
b
C
c

Production (m
3
/year) 250 730 3000 6000

pH 5.7 4.9 4.7 4.0 – 4.3
COD (mg/L) 1 200 – 10 266 5 200 14 150 9 240 – 17 900
BOD (mg/L) 130 – 5 320 2 500 8 100 5 540 – 11 340
TSS (mg/L) 385 – 5 200 522
e
1 060 1 960 – 5 800
TVS
d
(mg/L) 742 81 – 86 % of the TSS
Total N (kjeldahl) (mg/L) 12 – 93 61 48.2 74 – 260
Total P (mg/L) 23 25 5.5 16 to 68
a
Torrijos and Moletta (1998).
b
Vintage period, mean value after 24 h.
c
Extreme values.
d
TVS – Total volatile solids.
e
After primary sedimentation.
BREWERY AND WINERY 9

3.2.2. Treatment Processes
The criteria for selecting an anaerobic or an aerobic biological treatment are
identical in brewery and winery industries.
Like in the brewery industry, the winery wastewaters are characterized by their high
content on organic biodegradable compounds. In this case, the anaerobic technology is
the most economical bioprocess due to lower running costs for aeration and sludge
processing. However, as previously mentioned for the brewery case, the anaerobic

conversion is generally insufficient to attaint the effluent quality required for discharge
in surface waters. Therefore, the anaerobic treatment should be followed by an aerobic
system, if the option of co-treatment of the winery wastewaters in a (aerobic) municipal
wastewater treatment plant is not available. Despite such rule, in the case of small wine
industries where the minimization of investment costs is the key factor and only one
biological process may be considered, the option must be an aerobic process if the
objectives for effluent quality are high. Obviously, the financial burden of an aerobic
operation is not so heavy in the case of a low wastewater flow.
Organic matter is essentially in soluble form. Therefore, a static sedimentation unit
is not an option for significant concentration reduction. Besides, an important fraction of
the suspended matters is easily removed by settling (seeds, tartaric salts, filtration
earths). Another focal point is the removal of inorganic suspended solids from such type
of wastewaters because the abrasive solids used in precoated filters can damage
mechanical equipment. Furthermore, many biological processes face difficulties for
treating non-soluble wastewaters: a pre-treatment step using screening and/or
sedimentation is then mandatory.
The anaerobic process shows a very good reliability for winery wastewaters. The
COD/N/P ratio is appropriate for anaerobic bacteria and the seasonal activity is not a
problem for process start-up. The anaerobic digesters are generally heated to reach the
mesophilic range (but psychrophilic conditions are possible) and is advisable to measure
alkalinity routinely in order to avoid a sudden pH drop in one-stage processes. All
anaerobic technologies can be applied for treating winery wastewaters. Among them, two
of the most promising ones are granular UASB reactors and the anaerobic sequencing
batch reactor (aSBR). An interesting approach is reported by Keyser et al. (2003) who
evaluated three UASB reactors with the aim of tailoring granules for the treatment of
winery wastewater, a novel ecotechnological approach. One reactor was seeded with
granular sludge enriched with Enterobacter sakazakii and a 90 % COD removal at
hydraulic retention time of 24 h could be reached. This performance compares favourable
with a second reactor seeded with brewery granules that achieved 85 % COD removal
and with a third one seeded with municipal sludge, which showed problems and had

continuously to be re-seeded. Ruíz et al. (2002) operated an anaerobic sequencing batch
reactor at an organic loading rate, as COD, around 8.6 kg/(m
3
d) with soluble COD
(sCOD) removal efficiency greater than 98 %, hydraulic retention time of 2.2 d and a
specific organic loading rate, as COD/VSS (volatile suspended solids), of 0.96 g/(g d).
Anaerobic filters and completely mixed reactors are also used in the winery industry, but
fewer systems are under construction now.
As stated before, aerobic technologies are well suited for the depollution of
wastewaters from wineries, if their running costs are not decisive. Sequencing batch
reactors are becoming the most popular since Torrijos and Moletta (1997) used them to
10 A. G. BRITO ET AL
treat a winery wastewater and reported a 95 % sCOD elimination, and a nitrogen and
phosphorous removal of 50 % and 88 %, respectively. These results could be generalized
and the simplified automation and the possibility of coping with load fluctuations are
decisive SBR advantages. Nevertheless, other different designs are currently available.
Eusébio et al. (2004) have operated jet-loop reactors, Andreottola et al. (2005) performed
the treatment of a winery wastewater applying a two-stage fixed bed biofilm reactor, and
Coetzee et al. (2004) have implemented a pilot-scale rotating biological contactor. The
seasonal operation of wineries may be a problem for aerobic biological systems leading
to decreased sludge settleability, floc disintegration and increased solids in the treated
effluent (Chudoba and Pujol, 1996). Therefore, in order to work efficiently, even during
those temporary overloading periods, the plant has to be oversized. This strategy is rather
costly, because such a plant has to run below its nominal capacity during a major part of
the year.
In small wineries, simplified systems of low energy consumption – lagoons,
constructed wetlands, land spreading/irrigation – are also scenarios for effluent treatment
or polishing, but a landscape integration is sought and large areas of land should be
available (Bustamante et al., 2005). The feasibility of such approach depends on external
factors that restrain a generalized use, namely meteorological, hydrogeological, and soil

and biomass characteristics. Therefore, the engineering of a specific biological treatment
process for wineries wastewater, including the selection of ancillary equipment, should
be decided on a case by case basis, as stated by Rochard and Kerner (2004).
4. CASE STUDY 1: BREWING WASTEWATER TREATMENT
The brewery industry Unicer SA has in operation a UASB reactor (1600 m
3
)

for the
industrial wastewater treatment. The start-up of UASB reactors often rely on a massive
inoculation with biomass already in pellets/granules (Nollet et al., 2005), representing an
additional cost for
the brewery industry. Indeed, the Unicer SA reactor was inoculated
with granular sludge imported from a paper factory in Spain. A 70 % to 80 % COD
removal is generally recorded in the UASB process. In spite of such efficiency, the final
COD and ammonium nitrogen levels are above the threshold values prescribed by
legislation for wastewater discharge in surface waters. On the other hand, due to the
anaerobic digestion process, the carbon concentration in the UASB effluent is very low,
imposing difficulties on conventional post-denitrification processes. Therefore, as
depicted in Figure 5, several steps were performed. First, there was the formation of
anaerobic granules in a lab-scale UASB reactor using dispersed biomass as inoculum and
the industrial wastewater from Unicer SA as substrate. Second, the feasibility of SBR
technology for the post-treatment of the effluent from the UASB reactor was assessed.
For the post-treatment of the brewery wastewater, two different SBR strategies for
nitrogen removal were considered. One was based on an aerobic-anoxic sequence and the
other one comprised a pre-denitrification step, that is, an anoxic-aerobic-anoxic sequence.
In both tests, SBR performance and biological kinetics were evaluated.


BREWERY AND WINERY 11



Figure 5. Schematic diagram of the goals of the present chapter.
4.1. UASB Operation for the Formation of Biomass Granules
Non-aggregated biomass from an anaerobic digester used in the stabilization of
activated sludge was tested for granulation. The operational protocol was based on the
selection of aggregate-forming bacteria, mainly focused on the acetotrophic Methanothrix
spp, by favouring the wash-out of non-aggregated biomass (Hulshoff Pol, 1989). In order
to attain such objective, the loading rate was increased when acetate concentration was
lower than 50 mg/L, a value near the half saturation constant of Methanothrix spp.
During the first three months, the treated effluent was partly recycled to increase the
hydraulic load. The operating temperature in the UASB reactor was 35
o
C. The pH ranged
from 6.5 to 7.9.
Figures 6 and 7 show the operational conditions and results of the UASB reactor,
namely B
V
(volumetric organic load, organic matter mass concentration, as COD, per
time unit), COD (influent and effluent), and COD removal efficiency. The granular
activity sustained the application of high B
V
, up to 20 kg/(m
3
d), with average COD
removal efficiencies of 80 %. The objective of granulation process was successfully
achieved but a six month period of operation was necessary. The sedimentation velocity
of aggregated biomass attained 40 m/h to 50 m/h and the SVI (sludge volume index) was
10 mL/g. TS and TVS in granules amounted to 114 kg/m
3

and 87 kg/m
3
. Figure 8 shows
a SEM (scanning electron microscopy) picture of the granules, obtained at the end of
operation.
The feasibility of UASB reactor start-up based on an inoculation with non-
aggregated biomass was demonstrated for the treatment of brewery industry wastewaters,
concerning organic matter elimination. However, an amonification processes occurred,
NH
4
+
-N in the effluent ranging between 23 mg/L and 87 mg/L, while the influent NH
4
+
-N
was just 12 mg/L to 29 mg/L. Therefore, a further nitrogen removal process was
necessary in order to attain effluent thresholds for discharge into surface waters.




Effluent containing NH
4
+
-N higher
than the required level for discharge
into surface waters
Anaerobic
pre-treatment in a
full-scale UASB reactor

Lab UASB reactor to study the
formation of anaerobic granules
using a non-aggregated inoculum
Lab SBR for the post-treatment of the
brewery wastewater to provide a base
for the upgrading of Unicer SA
treatment system
UNICER SA
wastewater
12 A. G. BRITO ET AL
0
5
10
15
20
25
0 50 100 150 200 250
t /d
B
V
kg/(m
3
d)

Figure 6. Organic load applied to the UASB reactor.
0
1000
2000
3000
4000

0 50 100 150 200 250
t /d
COD
mg/L
0
20
40
60
80
100
COD
removal
efficiency
%

Figure 7. Results of UASB reactor operation along the operational time.
Legend: —— COD removal efficiency —æ— CODin —— CODout

Figure 8. SEM photograph of the biomass after granulation.
BREWERY AND WINERY 13

4.2. SBR Operation for the Post-Treatment of the Brewery Wastewater
The average composition of the UASB effluent collected at Unicer SA brewery is
shown in Table 3. The bench scale SBR was operated in the typical sequence of Fill,
React, Settle and Draw.
Two SBR operating strategies were tested during the present
study. Their main features are summarized in Figure 9. The SBR operational conditions
are described in Table 4. The biomass inoculum was a grab sample collected in a
municipal activated sludge plant of the extended aeration type (around 90 % of the
inoculum), supplemented with an inoculum of Alcaligenes denitrificans and nitrifying

microorganisms. The average MLVSS (mixed liquor volatile suspended solids concentra-
tion, mass per volume) during the experimental assays was 1690 mg/L. Solids sampling
represented the only biomass wastage carried out along the experimental work.
Therefore, the sludge age was rather long, being estimated as 37 d. The concentration of
nitrogen compounds in the treated effluent is depicted in Figure 10.
Table 3. Brewery wastewater composition after UASB pre-treatment
Parameter Range
pH 7.5 – 8.0
Total COD, tCOD (mg/L) 400 – 2 000
Soluble COD, sCOD (mg/L) < 470
Soluble organic carbon (C) (mg/L) 60 – 83
NH
4
+
-N (mg/L) 23 – 87
NO
2
–
-N (mg/L) 0 – 1.2
NO
3
–
-N (mg/L) 0 – 3
Soluble P (mg/L) 8 – 20
TSS (mg/L) 320 – 1440


Strategy 1: post-denitrification
Fill (0.5 h)


Aerobic (12 h)


Anoxic (12 h)

Settle (1 h)

Draw (0.5 h)


Fill (0.5 h)

Anoxic (2 h)

Strategy 2: pre-denitrification
Aerobic (4 h)

Anoxic (2 h)


Settle (1 h)

Draw (0.5 h)

Figure 9. SBR operational strategies.
14 A. G. BRITO ET AL
Table 4. SBR operating conditions
Strategy 1 Strategy 2
Reaction sequence


Aerobic (12 h)/
/Anoxic (12 h)
Anoxic (2 h)/Aerobic (4 h)/
/Anoxic (2 h)
Total cycle time (h) 26 10
Working volume (L) 2.9 1.7
Volumetric replacement (%) 60 30
HRT
a
(d) 1.9 1.2
DO (in the aerated phase) (mg/L) 3.7 2.8
N after Fill (mg/L) 30 – 45 20 – 28
Nitrogen (N) load per volume [kg/(m
3
d)] 0.040 0.086
Nitrogen (N) load per VSS
b
[kg/(kg d)] 0.024 0.051
a
HRT – Hydraulic retention time.
b
VSS – Volatile suspended solids.
Strategy 1 was characterized by the use of an aerated phase (dissolved oxygen
concentration, mass of O
2
per volume, DO = 3.7 mg/L) followed by an anoxic phase (see
Figure 11). Complete nitrification took place, during the aerated phase, ammonium and
nitrites being removed from the anaerobically pre-treated effluent. The maximum
observed SDR (specific denitrification rate, mass of nitrogen, N, per VSS per time) was
0.165 kg/(kg d). However, the nitrogen removal efficiency was 50 %, resulting in an

effluent NO
3
–
-N above 15 mg/L (the value prescribed by the legislation for discharge in
surface waters is 11 mg/L). Moreover, a nitrogen balance in the liquid phase showed that
NO
3
–
-N at the end of the aerobic phase (15 mg/L to 20 mg/L) was roughly 50 % lower
than the theoretically one expected according to reaction stoichiometry. Biomass yield
was not sufficient to fill this gap. Consequently, the data indicate the occurrence of a
significant denitrification process during the aerated phase.
0
10
20
30
40
50
60
70
80
0 20 40 60 80 100 120 140 160 180 200
t /d
NO
3
-
-N
mg/L
0
1

2
3
4
5
6
7
8
Average DO
mg / L
NH
4
+
-N
mg / L
NO
2
-
-N
mg / L
Strategy 1 Strategy 2

Figure 10. Long-term ammonium, nitrite and nitrate effluent concentrations and average DO levels in the
aerated phase. Legend: —— NH
4
+
-N —— NO
2
–
-N —— NO
3

–
-N —-— Average DO
BREWERY AND WINERY 15

0
10
20
30
40
50
0 5 10 15 20 25 30
t /h
ORP
mV
-150
-100
-50
0
50
100
150
NH
4
+
-N
mg/L
NO
2
-
-N

mg/L
NO
3
-
-N
mg/L
DO
mg/L
Aerobic
phase
Anoxic
phase

Figure 11. ORP (oxidation-reduction – redox – potential) values and DO, ammonium, nitrite and nitrate
nitrogen concentrations in the bulk liquid, along a typical SBR cycle, in strategy 1 (DO = 3.7 mg/L during the
aerated phase).
Legend: —— ORP —— DO —— NH
4
+
-N —— NO
2
–
-N —— NO
3
–
-N
As declared above, the maximum observed SDR was 0.165 kg/(kg d). An
explanation for such phenomenon relies on oxygen limitations within microbial flocs
providing the oxygen free conditions for heterotrophic denitrifying bacteria activity (van
Loosdrecht and Heijnen, 1993). Such hypothesis was tested setting a DO of 7 mg/L

during the aerated phase (Figure 12). In fact, at such high DO, denitrification did not
occur during the aerobic period, confirming that there was an oxygen limitation when the
bulk liquid DO was 3.7 mg/L.
The strategy 2 involved a pre-denitrification step and thereafter the aerated phase
(DO = 2.8 mg/L) and the anoxic phase. In Figure 13 the behavior of nitrogen compounds
and the ORP and DO profiles during a typical SBR cycle are shown.
The overall experimental results (Figure 10) demonstrated that the most appropriate
strategy for nitrogen removal in order to achieve the legal compliance for wastewater
discharge in surface waters was the anoxic-aerobic-anoxic sequence, with DO = 2.8 mg/L
in the aerated period, and a volumetric replacement of 30 % (strategy 2). Under such
conditions, the maximum observed SNRR (specific nitrogen removal rate) had the value
0.038 kg/(kg d), and NO
3
–
-N in the effluent was lower than 8 mg/L.
Thus, this strategy optimizes the energy requirements for aeration with an
appropriate effluent quality for discharge in surface waters. The denitrification during the
final anoxic phase (after the aerobic period) was practically meaningless in all runs.
16 A. G. BRITO ET AL
0
10
20
30
40
50
60
70
0 5 10 15 20 25 30
t /h
NH

4
+
-N
mg/L
NO
2
-
-N
mg/L
NO
3
-
-N
mg/L
0
50
100
150
200
250
300
sCOD
mg/L
Aerobic phase Anoxic phase

Figure 12. Ammonium, nitrite, nitrate and soluble COD in the bulk liquid along a typical SBR cycle in strategy
1 (during the aerated phase: DO = 7 mg/L).
Legend: —æ— NH
4
+

-N —— NO
2
–
-N —— NO
3
–
-N ——sCOD
The C/N ratio in the UASB effluent had an average value of 0.8 and carbon
requirements for complete nitrogen removal were not satisfied: the stoichiometric C/N
ratio, using an easily degradable carbon source like acetate, must be 1.25. The results
obtained when acetate was used to increase the mass C/N ratio to 1.3, during the anoxic
phase, leading to a complete nitrate removal, confirmed that the soluble carbon source
was limiting denitrification. Simultaneous nitrification and denitrification was detected
during the aerated phase at DO = 2.8 mg/L and 3.7 mg/L (Figures 11 and 13). On the
other hand, denitrification was inhibited during the aerated period when the bulk liquid
DO was raised to 7 mg/L (Figure 12). The redox potential was kept within the range
+100 mV to –240 mV, in response to oxygen concentration along each cycle. The ORP
provides information about the process regime and can be used to control the duration of
the denitrification phase (Demoulin et al., 1997). However, the typical breakpoint, the
“nitrate knee”, that appears in the ORP curve at NO
3
–
-N close to zero, could not be
observed (Figures 11 and 13). Due to the simultaneous nitrification-denitrification and
carbon limitations, nitrate was always present, even if at low concentrations, along the
whole operating cycle. An improvement of the biological floc settleability was noticed
along SBR operation. Soon after the start-up, flocs became larger and the SVI decreased
from 200 mL/g down to 115 mL/g. A concomitant decrease of TSS in the treated effluent
was observed, attaining only 30 mg/L at the end of the experimental period.
BREWERY AND WINERY 17


0
10
20
30
40
50
0246810
t /h
ORP
mV
sCOD
mg/L
-250
-150
-50
50
150
Anoxic
phase
Aerobic
phase
Anoxic
phase
NH
4
+
-N
mg/L
NO

3
-
-N
mg/L
DO
mg/L

Figure 13. Variation of ORP values, DO, soluble COD and nitrogen compounds concentration in the bulk
liquid along a typical SBR cycle, during strategy 2.
Legend: —— ORP —— DO —— sCOD —— NH
4
+
-N —— NO
3
–
-N
5. CASE STUDY 2: WINERY WASTEWATER TREATMENT
The start-up and optimization of the wastewater treatment process in the wine
industry are presented using the ACPB case study. The aimed optimization was based on
two operational strategies during the periods of high daily flows and organic loads.
5.1. Full-Scale Sequencing Batch Reactor Operation
A schematic diagram of the WWTP (wastewater treatment plant) is presented in
Figure 14.

Figure 14. Schematic diagram of the WWTP from ACPB: 1 – Sand remover; 2 – Equalization/neutralization
tank; 3 – Septic tank; 4 – Biological unit (SBR); 5 – Sludge thickener; 6 – River.
1
2
3
4

5
6
18 A. G. BRITO ET AL
A sand remover is located at the beginning of the WWTP in order to remove solid
materials, including diatomaceous earth, from the wastewater. If necessary, the correction
of the pH value is made in a 300 m
3
equalization tank, throughout CaCO
3
addition. Inside
the SBR, two superficial jet aerators with swing-arms are installed, with a capacity of
oxygenation of 5.5 kg/h each.
The SBR was inoculated with domestic sewage, with VSS = 540 mg/L. Nutrients
were supplied throughout the SBR feeding with domestic wastewater produced at ACPB.
The ratio BOD
5
/N/P was 100/10/0.4. The excess sludge from the SBR was conducted to a
gravity thickener. According to the working conditions, nine operational phases were
identified during the first year of operation of the SBR (Table 5). Each phase was either
related to a working period of the wine industry (with a typical effluent composition), or
to changes in the SBR operating conditions in order to increase the treatment efficiency.
In order to deal with the high effluent volumes generated during the vinification and
racking periods, the SBR was fed twice a day, representing a reduction of 50 % in the
reaction time. The time based SBR schedule is depicted in Table 6, describing the
operation with 1 and 2 cycles per day. Table 7 summarizes the general operating
conditions of the SBR, considering the operation with 1 cycle per day.
The pH values of the effluent were in the range between 7 and 8. Nevertheless,
during the vinification period, pH values of 3 were detected in the equalization tank. The
temperature of the effluent ranged between 15
o

C and 25
o
C. In general, high COD
removal efficiencies were detected, despite the B
V
changes (Figure 15) but during the
vinification and racking periods, B
V
increase led to a significant decrease in the COD
removal efficiency, due to oxygen limitations.
Table 5. Characterization of the different phases of WWTP operation
Operational
phase
Operational reactor
phases
Working period at the winery
Cycles
per phase
Cycles
per day
1 Start-up 65 1
2 Operation Washing operations and bottling 37 1
3 Operation Vinification 10 1
4 Operation Vinification and racking 29 1
5 Operation Bottling 24 1
6 Operation Second racking 42
1 → 2
7 1
st
sludge purge 26 2

8 2
nd
sludge purge 33 2
9
Biomass
recirculation
27 1
Table 6. Time based SBR schedule for operation with 1 and 2 cycles per day

Aerated fill Aerated react Settle Draw
1 cycle per day 0.5 h 21 h 2 h 0.5 h
2 cycles per day 0.5 h 10 h 1 h 0.5 h
BREWERY AND WINERY 19

Table 7. SBR general operating conditions (1 cycle per day)
Parameter Value
Working volume (m
3
) 150
Volumetric replacement (%) 17
HRT (d) 5.7
VSS (g/L) 2.5 – 4.5
B
V
[kg/(m
3
d)] 0.5 – 2.5
Applied specific load (as VSS) [kg/(kg d)] 0.26 – 0.57

In fact, B

V
, which was usually in the range between 0.5 kg/(m
3
d) and 1.5 kg/(m
3
d),
reached, in this period, the averaged value of 2.5 kg/(m
3
d). As a consequence, the
biomass concentration increased significantly and the oxygen supply was not enough to
fit the needs, resulting in tCOD values of 5000 mg/L in the discharge, despite the higher
COD removal rates. A wash-out of the biomass was observed for VSS higher than about
4.5 g/L, leading to an increase in the final effluent total COD and TSS. In fact, for VSS
higher than 4.5 g/L, the biomass exhibited a low sedimentation capability, due to the high
sludge age (45 d), leading to SVI > 120 mL/g. The results obtained showed that SVI
values should not exceed 80 mL/g, in order to maintain a good performance of the
biological reactor.
Towards the goal of increasing the SBR performance during the vinification and
racking periods and in order to account for the high daily flow and organic load of the
industrial effluent, two operational strategies were tested (Figure 16).
0
1
2
3
4
5
Operational phase
B
V
kg/(m

3
d)
0
10
20
30
40
50
60
70
80
90
100
COD
removal
efficiency
%
1 2 3 4 5 6 7 8 9

Figure 15. COD removal efficiency as a function of the applied volumetric load.
Legend: —æ— COD removal efficiency —— B
V

20 A. G. BRITO ET AL
0
20
40
60
80
100

120
0246810
SBR cycles
COD
removal
efficiency
%
0
0.5
1
1.5
2
2.5
B
V
kg/(m
3
d)

Figure 16. COD removal efficiency according to the applied organic load for different operational strategies (1
and 2 cycles per day).
Legend: —— Efficiency (1 cycle) —— Efficiency (2 cycles) —— B
V
(1 cycle) —— B
V
(2 cycles)
The first strategy, based on the operation of the SBR with two cycles per day
(resulting in a 50 % decrease in HRT, from 7.4 d to 3.7 d, and, therefore, in the
duplication of B
V

), was tested during the bottling period (average tCOD in the
equalization tank of 4000 mg/L). At this time, the winery wastewater comes, mainly,
from the washing operations and from the cooling processes, leading to high daily
wastewater flows. The second strategy was used when B
V
was high [above 1.5 kg/(m
3
d)]
and consisted of the recirculation of biomass from the SBR to the equalization tank, and
the use of an additional aeration system in both units, in order to provide the oxygen
needed for the organic matter biodegradation. The biomass recirculation to the
equalization tank and the aeration of the medium allowed the beginning of the
biodegradation processes at this stage, thus reducing the organic load applied to the SBR.
The results of the present study showed the suitability of a SBR designed on the
basis of averaged values of organic matter concentration and effluent flow, by changing
the operational strategy during the vinification and racking periods. In fact, during the
periods of high organic load (vinification and racking periods), the additional oxygen
supply led to a significant improvement in the WWTP performance, in terms of COD
elimination. During the rest of the year, the COD removal efficiency was always higher
than 90 % (Figure 16), despite the operation of the SBR with one or two cycles per day,
according to the industrial wastewater daily flow.
6. CONCLUSION
Brewery and winery industries are small and medium enterprises but with a
significant social and economic value. Therefore, their sustainability policy requires
wastewater treatment systems with the best performance and the fact is that well known
processes and technologies are available for such purpose. The experience obtained at
BREWERY AND WINERY 21

Unicer SA and ACPB demonstrated that the technological solutions are much site
specific – in their case, UASB and sequencing batch reactors were very appropriate –,

and highlighted that a good operation requires a bioengineering knowledge but is much a
continuous and endless effort in order to minimize costs maintaining the best quality and
service.
Acknowledgements
The authors are deeply thankful to Gerd Teunissen, Patrícia Moreira and Agostinha
Castro for their contribution during design, operation and analytical control of ACPB
WWTP and UASB lab-scale reactor. We also want to leave here our recognition to Luís
Melo for his pertinent research suggestions.
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